The objectives of the proposed study are to determine the influence of anthocyanin intake on fecal microbiota and to elucidate how obesity influences the effect of anthocyanin-induced changes on fecal microbiota. The goal of the Human Microbiome Project is to characterize microbial communities at different sites on the human body and identify relationships between the microbiome and human health. One initiative of the Human Microbiome Project is to elucidate the relationship between disease and changes in intestinal microbiota. The intestinal microbes have an important two-way relationship with nutrition in that foods consumed have an important impact on microbiota, and microbiota metabolize nutrients that pass into the colon, in some cases (such as flavonoids) producing active compounds that influence health and disease. Therefore, knowledge of the complex relationship between intestinal microbiota and nutrition will be important for elucidating the role of microbiota in human health. The goal of elucidating this relationship is in accord with the USDA National Program Priority to provide a scientific basis for dietary guidance for health promotion and disease prevention as well as the NIH focus on the microbiome. Berries are a rich source of flavonoids, especially the bright pigments called anthocyanins. Anthocyanins provide a variety of health benefits, which may include anti-cancer activity. Studies have suggested that specific anti-cancer effects may include decreased proliferation of neoplastic cells, signaling in apoptosis, suppression of inflammation, antioxidant activity, and suppression of DNA damage. It has been suggested that effects of anthocyanins and other polyhphenols may be attributable to metabolites as well as parent compounds. Only small amounts of anthocyanins and other polyphenols are absorbed from the small intestine, whereas most are broken down by colonic bacteria and enter systemic circulation as lower-weight metabolites. It is important to identify the relation between the parent compounds, their potentially cancer-preventive metabolites, and biomarkers for cancer. Answers to these questions can lead to tailored dietary advice for cancer prevention. The efficacy of berry flavonoids in reducing risk of cancer is influenced by metabolism, including how well flavonoids are absorbed, what metabolites are formed, and how quickly compounds are excreted. However, the metabolism of flavonoids is proving to be very complex. Studies with rodents suggest that flavonoid metabolism changes with adaptation to intake. Rats given a dose of flavonoids from grape extracts for two weeks had significantly different plasma kinetic curves than rats given a dose of flavonoids after a flavonoid-free diet. The mechanism for that adaptation remains completely unknown, but it may be related to changes in gut microflora with repeated exposure to flavonoids. Moreover, pilot data from our laboratory suggests that flavonoid absorption and/or metabolism is influenced by characteristics of metabolic syndrome. Individuals with low BMI and no risk factors for metabolic syndrome had significantly greater area under the curve than individuals with high BMI and tendencies toward metabolic syndrome (unpublished data, Figure 1). The mechanism is unknown, but again microbiota may play a role. Intestinal microbiota has been suggested to contribute to the development of metabolic syndrome and obesity, which is in turn associated with increased rates of cancer. Metabolomics offers a means for unraveling the mechanisms behind the complexities of flavonoid metabolism and for identifying metabolites with potent anti-cancer activity. Two biochemical markers were observed for berry intake, one marker known to be 3-hydroxyphenylacetic acid and the other yet unidentified. 3-Hydroxyphenylacetic acid was also observed to increase after intake of blackcurrant juice and red wine polyphenols and is likely a metabolite of microbial origin. Some of these colonic flavonoid metabolites are excreted into urine in significant amounts. For example, chlorogenic acids and flavan-3-ols produce a unique array of colonic metabolites which are excreted in substantial amounts in urine and correspond to as much as 29% of intake after coffee consumption and more than 70% of intake after tea consumption. Further, colonic metabolites are proving to be biologically active; for example, colonic polyphenol metabolites have been shown to help ameliorate protein glycation and neurodenegeration.